2381TRX-iFLP PKAalpha purification 2014 12 05

2014/12/1--12/5

2381 pT7A3-TRX-Tat-NLS-iFlp-6H 8L Nico21(DE3)plysN when OD600=2.0 use 0.2 mM IPTG RT induce overnight, 40g

add lysis buffer, freeze and thaw, then ultracentrifuge,

supernatant: load to yellow Ni column----elute ----diluted load to sp column--elute

pellet : use urea and  guanidine hydrochloride to dissolve, load to yellow Ni ----elute

lane1-4: 2381 protein eluted from sp column

lane5: 2381flowthrough from sp column

lane6-8: 2381 protein eluted from Ni column

PKAa

lane10: PKAa lysate

lane11: PKAa lysate SUPERNATANT purified with 75 ul Ni beads

lane12:PKAa lysate PELLET purified with 75 ul Ni beads